Target (protein): Type II site-specific deoxyribonuclease E.C Number: 3.1.21.4 Protein ID: WP_000781649.1 PDB ID: 1STX Organism: Escherichia coli Etiologic Risk Group (see link below): N/a Disease Information:
Escherichia Coli are bacteria that live in the intestines and are usually harmless but some of them can be pathogenic. They can cause diarrhea or infection outside of the intestinal tract. These types of bacteria are transmitted through contaminated water, food, or contact with another person or animal. The six types of E.Coli that cause this disease are Shiga toxin producing E.coli (STEC), Enterotoxigenic E. coli (ETEC), Enteropathogenic E. coli (EPEC), Enteroaggregative E. coli (EAEC), Enteroinvasive E. coli (EIEC) and Diffusely adherent E. coli (DAEC). Unfortunately, there is not a cure for E.Coli infections, it has to run its course and eventually get out of the body. To prevent from contracting en E.Coli infection maintain hands clean, cook meat thoroughly, wash vegetables, store foods separately, and get pasteurized drinks.
Links:
Essentiality of Type II site-specific deoxyribonuclease:
The Type II site-specific deoxyribonuclease is an enzyme that helps catalyze endonucleolytic cleavage of DNA to give specific double-stranded fragments with terminal 5'-phosphates. Links:
Reagents and Materials:
Relative activity in PCR mix (Taq DNA Polymerase buffer) is 10%. The PCR mix contained λDNA, primers, 10 mM Tris-HCl (pH 8.3, 20 °C), 50 mM KCl, 1.5 mM MgCl2, 200 μM dNTPs, 2.5 U Taq DNA polymerase. The mix was subjected to 25 amplification cycles. Activity in reaction buffer of Pwo SuperYield DNA Polymerase PCR Mix is 25%. When supplemented with GC-RICH Solution activity remains at 25%.
The Eco RV assay that contains all of these materials in it is worth $62.
Structure (PDB) Sequence ID: WP_000781649.1
Show pairwise alignment of your BLASTP search in NCBI:
Query Coverage: 100% Max % Identities: 100% % Positives: 100% Chain used for homology: A
Inhibitors: EDTA
Expression Information:
The target has been expressed in E.Coli. It is expressed predominatley in bacteria because it is a restriction modification system that helps protect the bacteria from foreign DNA that tries to inhibit it. Link:
*length of your protein in Amino Acids: 317 Molecular Weightof your protein in kiloDaltons using the Expasy ProtParam website: 35064.82 KDa Molar Extinction coefficient of your protein at 280 nm wavelength: 56505 TMpred graph Image (http://www.ch.embnet.org/software/TMPRED_form.html). Input your amino acid sequence to it.
Figure 2: TMPred output graph for Type II site-specific deoxyribonuclease *CDS Gene Sequence (paste as text only):
*GC% Content for gene: 7.886435% *CDS Gene Sequence (codon optimized) - copy from output of Primer Design Protocol (paste as text only): N/a *GC% Content for gene (codon optimized): N/a
E.C Number: 3.1.21.4
Protein ID: WP_000781649.1
PDB ID: 1STX
Organism: Escherichia coli
Etiologic Risk Group (see link below): N/a
Disease Information:
Escherichia Coli are bacteria that live in the intestines and are usually harmless but some of them can be pathogenic. They can cause diarrhea or infection outside of the intestinal tract. These types of bacteria are transmitted through contaminated water, food, or contact with another person or animal. The six types of E.Coli that cause this disease are Shiga toxin producing E.coli (STEC), Enterotoxigenic E. coli (ETEC), Enteropathogenic E. coli (EPEC), Enteroaggregative E. coli (EAEC), Enteroinvasive E. coli (EIEC) and Diffusely adherent E. coli (DAEC). Unfortunately, there is not a cure for E.Coli infections, it has to run its course and eventually get out of the body. To prevent from contracting en E.Coli infection maintain hands clean, cook meat thoroughly, wash vegetables, store foods separately, and get pasteurized drinks.
Links:
Link to TDR Targets page (if present): N/A
Essentiality of Type II site-specific deoxyribonuclease:
The Type II site-specific deoxyribonuclease is an enzyme that helps catalyze endonucleolytic cleavage of DNA to give specific double-stranded fragments with terminal 5'-phosphates.
Links:
Druggable Target: Not available
Link to BRENDA: http://www.brenda-enzymes.org/enzyme.php?ecno=3.1.21.4
Enzyme Assay Information: http://www.sciencedirect.com/science/article/pii/S002228360301297X
Eco RV assay of DNA using N6-methyladenine
http://www.sigmaaldrich.com/catalog/product/roche/ecorvro?lang=en®ion=US&cm_sp=Insite-_-prodRecCold_xviews-_-prodRecCold10-1
Reagents and Materials:
Relative activity in PCR mix (Taq DNA Polymerase buffer) is 10%. The PCR mix contained λDNA, primers, 10 mM Tris-HCl (pH 8.3, 20 °C), 50 mM KCl, 1.5 mM MgCl2, 200 μM dNTPs, 2.5 U Taq DNA polymerase. The mix was subjected to 25 amplification cycles. Activity in reaction buffer of Pwo SuperYield DNA Polymerase PCR Mix is 25%. When supplemented with GC-RICH Solution activity remains at 25%.
The Eco RV assay that contains all of these materials in it is worth $62.
Structure (PDB)
Sequence ID: WP_000781649.1
Show pairwise alignment of your BLASTP search in NCBI:
Query Coverage: 100%
Max % Identities: 100%
% Positives: 100%
Chain used for homology: A
Inhibitors: EDTA
Expression Information:
The target has been expressed in E.Coli. It is expressed predominatley in bacteria because it is a restriction modification system that helps protect the bacteria from foreign DNA that tries to inhibit it.
Link:
Purification Method: Heparin-Sepharose column chromatography
Image of Protein:
Figure 1: Structure of the K38A mutant of EcoRV bound to cognate DNA and Mn2
*Amino Acid Sequence (paste as text only - not as screenshot or as 'code'):
<span style="background-color: #ffffff; color: #626262;"> 1__0__ 2__0__ 3__0__ 4__0__ 5__0__ 6__0__ MNNQNDYIEA AQQIIASLGL PRAQQNERSA LCLLALLNLT PGKAWADAEN PLVGITPIMN 7__0__ 8__0__ 9__0__ 10__0__ 11__0__ 12__0__ WVREHYGKVY APNTRETFRR QSMHQFCAAG VALYNPDKPD RPVNSPKAVY QIEPAALSML 13__0__ 14__0__ 15__0__ 16__0__ 17__0__ 18__0__ RTFGSPAWHD SLATYLAERE TLVTRYAKER EQNRIPVEIA AGQQITLSPG EHSELIRAII 19__0__ 20__0__ 21__0__ 22__0__ 23__0__ 24__0__ EDFAPRFAPG SVLVYAGDTG EKWGYFDAPL LAGLGVDVDS HGKMPDVVLH FTAKNWLLLV 25__0__ 26__0__ 27__0__ 28__0__ 29__0__ 30__0__ ESVTSHGPVD GKRHAELARL FAGSTAGLVY VTAFPNRSIM GRYLGEIAWE TEVWVADAPS 31__0__ HLIHFNGVRF LGPYSTE </span>*length of your protein in Amino Acids: 317Molecular Weight of your protein in kiloDaltons using the Expasy ProtParam website: 35064.82 KDa
Molar Extinction coefficient of your protein at 280 nm wavelength: 56505
TMpred graph Image (http://www.ch.embnet.org/software/TMPRED_form.html). Input your amino acid sequence to it.
Figure 2: TMPred output graph for Type II site-specific deoxyribonuclease
*CDS Gene Sequence (paste as text only):
*GC% Content for gene: 7.886435%
*CDS Gene Sequence (codon optimized) - copy from output of Primer Design Protocol (paste as text only): N/a
*GC% Content for gene (codon optimized): N/a